1. Field of the Invention
The present invention is directed to the field of delivery systems for delivering pharmaceutical agents to a target tissue. In particular, the present invention is directed to a delivery system where hyaluronidase is located on the surface of nanoparticles to enhance the nanoparticle diffusion in the tissue and/or a relatively low density of second polyethylene glycol layer is located on the surface of nanoparticles to prolong nanoparticle circulation time in an animal or human body.
2. Description of the Related Technology
Nanoparticles alter the pharmacokinetics and toxicity of encapsulated drugs, leading to improved efficacy and reduced side effects of the drugs. Particularly in the therapeutic area of tumors, leaky vasculature and impaired lymphatic drainage of tumors allow the nanoparticles to enter and stay in the tumors. A few nanomedicines have been approved by the U.S. Food and Drug Administration (FDA) for cancer therapy, including albumin-bound paclitaxel for metastatic breast cancer. Some others are in clinical trials, for example Methoxy-PEG-poly(D,L-lactide) taxol (Genexol®-PM) has been approved in South Korea to treat metastatic breast cancer, though is still in phase III trials in U.S. (NCT00876486).
In spite of these limited successes, there are two major challenges in applying nanomedicines in cancer therapy. One is the quick clearance of synthetic nanoparticles by host immune systems before the nanoparticles can reach the tumor sites. The other challenge is the low interstitial diffusion of nanoparticles after entering perivascular areas in solid tumors. Released drugs only penetrate a few layers of tumor cells and are thus inaccessible to hypoxic tumor cells which are usually resistant to chemo- and radiotherapies.
Some progress has been made to address these two challenges. It is known that, through adsorption of opsonin proteins on synthetic nanoparticles, the synthetic nanoparticles become visible to phagocytic cells, resulting in quick clearance of the nanoparticles from blood circulation. Polyethylene glycol (PEG) and some zwitterionic polymers are known to prevent protein adsorption. Nanoparticles (NPs) coated with these polymers have extended circulation times and improved bioavailability of one or more encapsulated pharmaceutical agent. One example is the FDA-approved Doxil which are PEG-coated liposomes with encapsulated doxorubicin.
The low interstitial diffusion of nanoparticles in tumors is mostly due to the elevated density of cells and extracellular matrix (ECM), the high interstitial pressure and heterogeneous vasculature in the tumors. Some small-sized nanoparticles displayed improved diffusion within tumors. However, this strategy is often not practical as small-sized drug nanoparticles are difficult to fabricate. Further, even small nanoparticles still face a low diffusion in tumors as antibodies a few nanometers in size still exhibit the diffusion problem.
Some electronic device-assisted methods have also been developed to enhance nanoparticle diffusion in tumors, such as by reducing interstitial fluid pressure, magnetic field-assisted penetration, and generation of acoustic cavitation by ultrasound. These strategies may be effective for some specific types of tumors, but are not effective in treating metastatic tumors.
A potentially promising approach is the use of a tumor penetrating peptide to enhance diffusion of nanoparticles in the tumor. The peptide has a short motif targeting αv integrins that are highly expressed in tumor vasculature, and can be proteolytically cleaved in tumors to bind neuropilin-1. This binding increases the penetration of antibodies and nanoparticles in tumors though the actual mechanism is not clear.
Gou et al. “Development of theranostic nanoparticles with the ability to break extracellular cellular matrix for enhanced nanoparticle delivery,” NanoTech Conference and Expo 2012, Jun. 18-21, 2012, (Abstract) discloses nanoparticles conjugated with two extracellular matrix proteases, collagenase and/or hyaluronidase. These proteases can temporarily break down the extracellular matrix and open microscopic channels, allowing nanoparticles to spread in the tumor. The proteases were conjugated on polymer-coated quantum dots. After incubation with the 4T1 mammary breast cancer cell line, such nanoparticles were found capable of binding to the cells with intensive binding in the invasive edge of tumor cell clones. It was further found that collagenase displays even better improvement on nanoparticle diffusion in tumors, in comparison with hyaluronidase.
US 2008/0267876 provides a delivery system comprising a polymer-based nanoparticle (NP), and a linker comprising a first portion non-covalently anchored to the nanoparticle, where at least part of the first portion comprises a hydrophobic/lipophilic segment embedded in the nanoparticle, and a second portion comprising a maleimide compound exposed at the outer surface of the nanoparticle. In accordance with one embodiment, the delivery system comprises one or more targeting agents (e.g., antibody or ligand), each covalently bound to the maleimide compound. In accordance with yet another embodiment, the delivery system comprises a drug.
US 2005/0227911 provides hydrophilic dispersions comprising complexes consisting essentially of nanosized particles of a macromolecule wrapped in an amphiphilic polymer such that non-valent bonds are formed between the macromolecule and the amphiphilic polymer. The macromolecules may be a naturally-occurring, synthetic or recombinant polypeptide, protein, polysaccharide or polynucleotide, and the amphiphilic polymer is a polysaccharide or a modified polysaccharide such as starch, chitosan or an alginate. The protein may be hyaluronidase, among a long list of options.
US 2013/0337066 provides a nanoparticle comprising an inner core including a non-cellular material, and an outer surface comprising a membrane derived from a cell or a virus. The nanoparticle may be used in medicament delivery systems where pharmaceutical compositions comprise the nanoparticles. Immunogenic compositions comprising the nanoparticle can be used for eliciting an immune response, and for treating or preventing diseases or conditions, such as neoplasm or cancer, or diseases or conditions associated with cell membrane inserting toxin.
U.S. Pat. No. 7,767,429 discloses soluble neutral active Hyaluronidase Glycoproteins (sHASEGP's) and their use to facilitate administration of other molecules or alleviate glycosaminoglycan associated pathologies. The soluble, neutral active sHASEGP's include asparagine-linked sugar moieties required for a functional neutral active hyaluronidase domain and/or modified amino-terminal leader peptides that enhance secretion of sHASEGP's.
US 2011/0097277 discloses a nanoparticle comprising a core and a surface having a plurality of zwitterionic polymers grafted thereto or grafted therefrom. The core comprises a metal, a metal oxide, a ceramic, a synthetic polymer, a natural polymer, silicon dioxide, a crystal, a semiconductor material, a hydrogel, a liposome, a micelle, or a carbon-based material. The zwitterionic polymer has the formula: PB-(L1-N+(Ra)(Rb)-L2-A(=O)OM)n(X−)n, wherein PB is a polymer backbone having n pendant groups L1-N+(Ra)(Rb)-L2-A(=O)OM); N+is a cationic center; Ra and Rb are independently optional as necessary to provide a cationic center and independently selected from alkyl and aryl; A(=O)OM is the anionic center, wherein A is C, S, SO, P, or PO, and wherein M is a counterion; L1 is a linker that covalently couples the cationic center to the polymer backbone; L2 is a linker that covalently couples the cationic center to the anionic center; X− is the counter ion associated with the cationic center; and n is an integer from 1 to about 10,000.
U.S. Pat. No. 6,007,845 discloses nanoparticles that are not rapidly cleared from the blood stream by the macrophages of the reticuloendothelial system. The nanoparticles have a core of a multiblock copolymer formed by covalently linking a multifunctional compound with one or more hydrophobic polymers and one or more hydrophilic polymers, and contain a biologically active material. The terminal hydroxyl group of the poly(alkylene glycol) can be used to covalently attach onto the surface of the particles biologically active molecules, including antibodies targeted to specific cells or organs, or molecules affecting the charge, lipophilicity or hydrophilicity of the nanoparticle. The nanoparticles have a prolonged half-life in the blood compared to nanoparticles not containing poly(alkylene glycol) moieties on the surface.
U.S. Pat. No. 5,543,158 discloses nanoparticles that are not rapidly cleared from the blood stream by the macrophages of the reticuloendothelial system, and that can be modified to achieve variable release rates or to target specific cells or organs. The nanoparticles have a biodegradable solid core containing a biologically active material and poly(alkylene glycol) moieties on the surface. The terminal hydroxyl group of the poly(alkylene glycol) can be used to covalently attach onto the surface of the nanoparticles biologically active molecules, including antibodies targeted to specific cells or organs, or molecules affecting the charge, lipophilicity or hydrophilicity of the particle. The surface of the nanoparticle can also be modified by attaching biodegradable polymers of the same structure as those forming the core of the particles.
The present invention provides a therapeutic delivery system that transports hyaluronidase to a tissue, where the hyaluronidase digests the hyaluronan (or hyaluronic acid) in the extracellular matrix of the tissue to improve diffusion of nanoparticles or nanomedicine into the tissue. The delivery system differs from the prior system of Gou et al. because the present invention uses, for example, therapeutic-encapsulating organic nanoparticles that are biocompatible and degradable, while the system of Gou et al. uses inorganic nanoparticles, which have no potential to encapsulate therapeutics. Gou et al. does not disclose detailed procedures about the fabrication of the inorganic nanoparticles. However, it is well known that the methods of fabricating inorganic and organic nanoparticles are dramatically different, and are not interchangeable. In addition, the present invention achieved different and even unexpected results, by using hyaluronidase to significantly enhance organic nanoparticle diffusion and penetration through a matrix including hyaluronan, a core value of the present invention. The present invention also demonstrated i) that hyaluronidase on nanoparticle surfaces is dramatically more efficient than free/or called unconjugated hyaluronidase in assisting nanoparticle diffusion/penetration. ii) the surface modification of hyaluronidase has minor or negligible effect to increase nanoparticle binding to cells. These results were not disclosed in Gou et al.